DNA from five lines of transformed bladder epithelial cells derived from cultures of primary cells that had been treated with 7,12‐dimethylbenz[a]anthracene (DMBA) can transform NIH 3T3 mouse fibroblasts in DNA transfection experiments. Southern analysis of DNA from NIH 3T3 primary and secondary transformants established that four of the DMBA‐transformed cell lines contained activated cellular Ki‐ras, while the remaining cell line contained a transforming gene that is unrelated to Ki‐ras, N‐ras, and Ha‐ras. The point mutations responsible for Ki‐ras activation were detected using oligonucleotide probes following selective amplification of Ki‐ras specific sequences using the polymerase chain reaction. The results showed that activation of Ki‐ras invariably involved a GC → AT transition mutation of the first position of codon 12. Surprisingly, a Ki‐ras gene that was activated by a GC → AT transition mutation at the same position was also detected in a single transformed bladder urothelial cell line derived from control cultures of mouse bladder cells. Together, our results indicate that Ki‐ras activation in the DMBA‐transformed bladder cell lines may not be a direct consequence of interaction of activated DMBA metabolites with the Ki‐ras gene.
- Key words
- Ki‐ras genes